Imaging method and system using substrate functionalization

Abstract

An imaging system for analyzing fluorescent molecules in a sample, including a confocal microscope device, has a support in contact with at least a portion of the sample. In the system, the support surface in contact with the sample is functionalized so as to reduce the observation volume of the microscope on the surface in the axial direction. The present disclosure also relates to various uses of such a system as well as to a method for analyzing fluorescent molecules in a sample, the method being implemented by such a system.

The invention claimed is: 
     
       1. Imaging system for analyzing fluorescent molecules in a sample, the system comprising said sample and a confocal or large-field microscopy device comprising a support in contact with at least part of said sample, a surface of the support in contact with said sample comprises luminescent nano-objects able to give rise to an excitation of the fluorescent molecules present in the sample by non-radiative energy transfer of a Forster type (FRET) when said nano-objects are previously excited. 
     
     
       2. Imaging system according to  claim 1 , wherein the luminescent nano-objects are previously excited by a light radiation or an electrical excitation. 
     
     
       3. Imaging system according to  claim 2 , wherein the luminescent nano-objects are previously excited by a laser light radiation emitting in blue. 
     
     
       4. Imaging system according to  claim 1 , wherein the luminescent nano-objects are luminescent semiconductor nano-crystals. 
     
     
       5. Imaging system according to  claim 1 , further comprising at least two types of different luminescent nano-objects arranged so that energies thereof are transferred non-radiatively in a direction of molecules present at and close to said surface and of different natures. 
     
     
       6. Imaging system according to  claim 1 , wherein the microscopy device is provided with illumination means able to emit a light beam, and provided with a focusing means arranged firstly so as to direct said light beam to the sample and secondly so as to collect fluorescence light intensity emitted by the fluorescent molecules present in the sample. 
     
     
       7. Imaging system according to  claim 1 , wherein the microscopy device is provided with a spatial filtering means able to create an observation volume encompassing the luminescent nano-objects present at said surface of the support and the fluorescent molecules present in the sample. 
     
     
       8. Use of an imaging system according to  claim 1  for producing a fluorescence correlation spectroscopy system, having the confocal microscopy device being provided with an illumination means able to emit an excitation light beam in a direction of the sample. 
     
     
       9. Use of a large-field imaging system according to  claim 1  for determining positions of points of contact between a living cell and the support of said system. 
     
     
       10. The imaging system according to  claim 1 , wherein said device is a confocal microscopy device,
 said nano-objects are excited, and 
 fluorescence light intensity emitted by fluorescent molecules present in the sample are collected. 
 
     
     
       11. The imaging system according to  claim 10 , wherein the luminescent nano-objects are previously excited by light radiation or an electrical excitation. 
     
     
       12. The imaging system according to  claim 10 , wherein the luminescent nano-objects are previously excited by laser light radiation emitting in blue.